Jianing Yang

High sensitivity PCR assay in plastic micro reactors

Small volume operation and rapid thermal cycling have been subjects of numerous reports in micro reactor chip development. Sensitivity aspects of the micro PCR reactor have not been studied in detail, however, despite the fact that detection of rare targets or trace genomic material from clinical and/or environmental samples has been a great challenge for microfluidic devices. In this study, a serpentine shaped thin (0.75 mm) polycarbonate plastic PCR micro reactor was designed, constructed, and tested for not only its rapid operation and efficiency, but also its detection sensitivity and specificity, in amplification of Escherichia coli (E. coli) K12-specific gene fragment. At a template concentration as low as 10 E. coli cells (equivalent to 50 fg genomic DNA), a K12-specific gene product (221 bp) was adequately amplified with a total of 30 cycles in 30 min. Sensitivity of the PCR micro reactor was demonstrated with its ability to amplify K12-specific gene from 10 cells in the presence of 2% blood. Specificity of the polycarbonate PCR micro reactor was also proven through multiplex PCR and/or amplification of different pathogen-specific genes. This is, to our knowledge, the first systematic study of assay sensitivity and specificity performed in plastic, disposable micro PCR devices.

FULLY INTEGRATED MICROFLUIDIC BIOCHIPS FOR DNA ANALYSIS

Microfluidics-based biochip devices are developed to perform DNA analysis from complex biological sample solutions. Microfluidic mixers, valves, pumps, channels, chambers, heaters, and DNA microarray sensor are integrated to perform magnetic bead-based rare cell capture, cell preconcentration and purification, cell lysis, polymerase chain reaction, DNA hybridization and electrochemical detection in a single, fully automated biochip device. No external pressure sources, mechanical pumps, or valves are necessary for fluid manipulation, thus eliminating sample contamination and simplifying device operation. Pathogenic bacteria detection and single-nucleotide polymorphism analysis directly from blood are demonstrated. The device with capability of on-chip sample preparation and DNA detection provides a cost-effective solution to direct sample-to-answer genetic analysis, and thus has potential impact in the fields of point-of-care genetic analysis and disease diagnosis.

Acoustic microstreaming for biological sample mixing enhancement

A mixing technique based on bubble-induced acoustic microstreaming principle was developed. A mixer consists of a piezoelectric disk that is attached to a reaction chamber, which has a set of air bubbles with desirable size trapped in the. solution. Fluidic experiments showed that air bubbles resting on a solid surface and set into vibration by the sound field generated circulatory flows, resulting in global convection flows and thus rapid mixing. The time to fully mix a 100 /spl mu/L chamber is significantly reduced from hours (diffusion-only) to tens of seconds. CFD modeling showed that the induced flowfield and thus degree of mixing strongly depends on bubble positions. Immuno-magnetic cell capture experiments showed acoustic microstreaming provided efficient mixing of bacterial cell (E. coli K12) matrix suspended in blood with magnetic capture beads, resulting highly effective immuno-magnetic cell capture. Bacterial viability assay experiments showed that acoustic microstreaming has a relatively low shear strain field since the blood cells and bacteria remained intact after mixing: Acoustic microstreaming has many advantages over most existing chamber micromixing techniques, including simple apparatus, ease of implementation, low power consumption (2 mW), and low cost.

Fully Integrated MCM Microfluidic System for DNA Analysis

A fully integrated and self-contained microfluidic multi-chip modular (MCM) device having sample-to-answer capability was developed. The device performs complete genetic analysis, including sample preparation (i.e., target cell capture, cell purification/concentration, cell lysis, DNA amplification) and microarray detection.